Fig. 1. Image segmentation diagram

Fig. 2. Flow chart of the simulated annealing algorithm

Fig. 3. Flow chart of static autofocus

Fig. 4. Evaluation function diagram of ideal focus/defocus images

Fig. 5. Flow chart of defocus detection

Fig. 6. Flow chart of dynamic real-time focusing

Fig. 7. Composition diagram of microscope auto-focusing experimental equipment

Fig. 8. Auto-focusing experimental equipment of microscope

Fig. 9. Focused images. (a) Pipeline model; (b) oil-water interface; (c) small blisters in the oil; (d) oil-water emulsion 1; (e) big blisters in the oil ; (f) oil-water emulsion 2

Fig. 10. Curve comparison of different methods. (a) Pipeline model; (b) oil-water interface; (c) small blisters in the oil; (d) oil-water emulsion 1; (e) big blisters in the oil ; (f) oil-water emulsion 2

Fig. 11. Comparison of image evaluation function curves. (a) Pipeline model; (b) oil-water interface; (c) small blisters in the oil; (d) oil-water emulsion 1; (e) big blisters in the oil ; (f) oil-water emulsion 2

Fig. 12. Simulated local extremum function curve. (a) Image group 1; (b) image group 2; (c) image group 3; (d) image group 4

Fig. 13. Focus/defocus evaluation curve and defocus detection curve of the same images

Fig. 14. Defocus level curve of dynamic observation images

Fig. 15. Real-time sample defocus amount average value curve